PROJECT SUMMARY The goal of this project is to define the role of peroxisome proliferator-activated receptor (PPAR)3 activation in neointima formation. Neointima formation occurs frequently after angioplasty and causes significant morbidity; vascular smooth muscle cells (SMCs) are key cells during neointima formation. We will study the effects of two clinically available agents on SMC biology and neointima formation. PPAR3 is a ligand-activated nuclear receptor that has been shown to have beneficial effects on vascular disorders. We will compare the effects of two agents: pioglitazone (activates PPAR3 only) and bexarotene (an RXR agonist which activates PPAR3, PPAR1, PPAR4, LXR, and FXR). Our hypothesis is that PPAR3 activation specifically in smooth muscle cells (SMC) will reduce neointima formation by decreasing resident SMC migration and proliferation as well as SMC-derived chemokine production and subsequent recruitment of bone marrow-derived cells. We believe both pioglitazone and bexarotene will be effective but bexarotene may be more effective due to activation of other nuclear receptors. In Aim One, we will compare the effects of pioglitazone to bexarotene on SMCs. We will measure changes in proliferation, cytokine production, and smooth muscle gene expression. In Aim Two, we will determine if the agents affect levels of microRNAs crucial to maintaining SMC phenotype, such as miR- 143, miR-145, and miR-221. In Aim Three, we will examine the effects of the agents in vivo during femoral artery wire injury. To track recruitment of bone-marrow derived cells to the site of arterial injury, all mice will receive bone marrow transplants from a GFP positive donor. After wire injury, mice will be analyzed at multiple time points. Along with neointima size, we will measure production of chemokines (IL-6, MCP-1, SDF-11, and KC), recruitment of bone marrow-derived cells and macrophages, and cellular proliferation. We also plan to study the role of PPAR3 activation specifically in smooth muscle cells during neointima formation. Using an inducible tissue-specific knockout model, we will deplete PPAR3 in smooth muscle cells after mice have received bone marrow transplants from GFP positive donors. Inducible PPAR3 knockout mice and control mice will receive therapy with either pioglitazone, bexarotene or control and be subjected to femoral artery wire injury. At multiple time points, neointima size will again be measured. All mice used in Aim 3B will have the R26R reporter allele in which smooth muscle cells are labeled with 2-galactosidase. Since we can measure both bone marrow derived and resident SMCs, we will determine the relative contribution each cell type makes to the neointima. We will also be able to determine if PPAR3 specifically in smooth muscle cells mediates the effects of bexarotene or pioglitazone.